Over the last 10 years, the lysosome-mediated degradation pathway macroautophagy has gained prominence in the field of adult onset neurodegeneration. Macroautophagy is an essential cellular pathway responsible for the elimination of cytosolic proteins, lipids and organelles. It is the most conserved of the three forms of autophagy, and its importance in mammals can be seen in a wide array of processes from the earliest stages of development to the maintenance of the aging brain. Although neurons were among the earliest cell types to be examined for macroautophagy, subsequent biochemical and genetic studies in liver and yeast caused the interest in neuronal macroautophagy to wane. Nonetheless, as the importance of macroautophagy in neurodegenerative diseases continues to grow, it is clear that we must refocus our attention on neural macroautophagy if we are truly to understand how this essential pathway impacts the adult brain. In this proposal, we will examine two key questions that will define how fundamental macroautophagic events are relevant in the healthy and diseased adult brain. In Aim 1, we will define how membrane cholesterol impacts macroautophagy, and how disruptions in cholesterol homeostasis may lead to dysfunctional autophagy in the mammalian brain. Then in Aim 2, we will determine if aggregation-prone proteins are indeed eliminated by macroautophagy in the adult brain, and establish the minimal components essential for this process. By answering these questions, we will not only further our mechanistic understanding of key macroautophagic processes, but we will also understand them in a context that will further our understanding of how macroautophagy can impact neurodegenerative disease.